| Objective:To observe the effects of Yiqi Tongyang Zhuyu Shengxin Recipe(YQT YF)on the pathological morphology of ischemic myocardium,microvascular endothelial cells and angiogenesis,the expression of VEGF,bFGF,Ang-land Shh signaling pathway related factors(Shh,Patched-1,Smo,Gli-1)in myocardial infarction border area of rats with myocardial ischemia-reperfusion injury,the mechanism of action on cardiac angiogenesis in MIRI rats.Methods:A total of 120 SPF male SD rats were divided into Sham group(n=20)and model group(n=100)according to random number table.Acute MIRI model was established by ligation of the left anterior descending coronary artery.According to the stratification results,they were randomly divided into model group,tanshinone ⅡA group,YQTYF group,Cyclopamine group(a specific inhibitor of Shh signaling pathway),and YQT YF+Cyclopamine group.After 7 days,the rats were sacrificed in groups,and the microvascular endothelial structure and microangiogenesis in the ischemic myocardium at the edge of myocardial infarction were observed by TEM.Pathological changes of ischemic myocardium at the edge of myocardial infarction in rats were observed by Hematoxylin Eosin staining.The microvessel density and the expression levels of angiogenic factors VEGF,bFGF and Ang-1 in the ischemic myocardium at the edge of myocardial infarction were observed by immunohistochemistry.The protein expression levels of Shh,Patched-1,Smo,Gli-1 in Shh signaling pathway were observed by Western Blot.Results:1.The results of transmission electron microscopy showed that:The basal membrane of vascular endothelial cells was intact and there was no obvious edema of mitochondria in the sham operation group.In the model group,there was obvious ischemic lesion,vascular endothelial cell swelling and mitochondria swelling.Compared with the model group,the basal membrane of vascular endothelial cells in tanshinone IIA group was intact,and the mitochondrial swelling was reduced.The ischemic damage in the YQTYF group was significantly improved,the vascular endothelial cell membrane was basically intact,and the mitochondrial swelling was reduced.In the Cyclopamine group,there were significant ischemic changes,the microvascular endothelial cells were seriously damaged and the surrounding space was enlarged,the microvessel was wrinkled and mitochondria were swollen.Vascular endothelial cells,incomplete cell membrane and mitochondria of the YQTYF+Cyclopamine group were swollen.2.The results of HE staining showed that:The myocardial cells in the edge area of myocardial infarction in the sham operation group were basically normal.The myocardial cells of model group showed obvious ischemic damage.Compared with the model group,the ischemic myocardial cell injury was significantly reduced in tanshinone IIA group and YQTYF group,while the ischemic myocardial cell injury was significantly reduced in the Cyclopamine group,while the ischemic myocardial cell injury was slightly reduced in the YQTYF+Cyclopamine group compared with the model group.3.The results of CD31 labeled MVD showed that:Compared with the sham operation group,the MVD of ischemic myocardi altissue in the infarction edge area of the model group was significantly increased(P<0.01);Compared with model group,the expression of MVD in tanshinone IIA group and YQTYF group was increased,while the expression of MVD in Cyclopamine group and YQTYF+Cyclopamine group was significantly down-regulated,with statistical significance(P<0.05 or P<0.01).Compared with tanshinone IIA group,there was no significant difference in MVD expression in the YQTYF group(P>0.05),while the down-regulation of MVD expression was significant in the Cyclopamine group and YQTYF+Cyclopamine group(P<0.01).Compared with Cyclopamine group,the expression of MVD in YQTYF,YQTYF+Cyclopamine group was up-regulated(P<0.05).4.The expression results of angiogenic factors VEGF,bFGF and Ang-1 showed that:Compared with the sham operation grou p,the protein expressions of VEGF,bFGF and Ang-1 in the isch emic area of rats in the model group were increased(P<0.0 5or P<0.01).Compared with model group,the protein expressions of VE GF,bFGF and Ang-1 in tanshinone IIA group and YQTYF were significantly increased,while the protein expressions of VEGF,bF GF and Ang-1 in Cyclopamine group and YQTYF+Cyclopamine g roup were decreased(P<0.05 or P<0.01).Compared with tanshinoneⅡA group,there was no significant difference in YQTYF group(P>0.05),but the expressions of VEGF,bFGF and Ang-1 were significantly down regulated in YQTYF group and YQTYF+Cycl opamine group(P<0.05 or P<0.0 1);The expressions of VEGF,bFG F and Ang-1 in YQTYF and YQTYF+Cyclopamine groups were h igher than those in Cyclopamine group(P<0.0 5 or P<0.0 1).5.The expression results of Shh,Patched-1,Smo,and Gli-1 sh owed that:Compared with the sham operation group,the protein e xpressions of Shh,Patched-1,Smo,and Gli-1 in the model group we re significantly down-regulated(P<0.01);Compared with the model group,the protein expressions of Shh,Patched-1,Smo,and Gli-1 i n tanshinone IIA group and YQTYF group were significantly upregulated,while the protein expressions of Shh,Patched-1,Smo,and Gli-1 in the group of Cyclopamine group and YQT YF+Cy clopa mine group were down-regulated(P<0.05 or P<0.01).Compared with tanshinone IIA group,the protein expressions of Shh,Patched-1,S mo,and Gli-lin the YQT YF group and YQ TYF+Cy cl opamine grou p were significantly down-regulated,and the differences were stat istically significant(P<0.01).Compared with Cyclopamine group,t he protein expressions of Shh,Patched-1,Smo,and Gli-1 in YQT YF,YQTYF+Cycl op amine group were up-regulated(P<0.05 or P<0.0Conclusion:1.YQTYF can improve the pathological morphology of ischemic myocardium and the ultrastructure of vascular endothelial cells,promote cardiac angiogenesis,and has a certain protective effect on myocardial ischemia-reperfusion injury model rats.2.YQTYF can up regulate the expression of VEGF,bFGF,Ang-1,Shh,P at ched-1,Smo,Gli-1,which are the main factors of Shh signaling pathway in the ischemic myocardium of the border area of myocardial infarction,increase the synthesis and secretion of these protein factors,cause angiogenesis in the ischemic myocardium of the border area of myocardial infarction,so as to establish a good collateral circulation.3.YQTYF may promote the expression of VEGF,bFGF and Ang-1 protein by activating Shh signaling pathway to achieve angiogenesis. |
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